NER: Biological Nanomachines: Assembly and Function of Protein-DNA Nanostructures at the Single-Molecule Level

NER：生物纳米机器：单分子水平上蛋白质-DNA 纳米结构的组装和功能

• 批准号: 0103556
• 负责人: Stephen Kowalczykowski
• 金额: $ 10万
• 依托单位: University of California-Davis
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-07-01 至 2002-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0103556&HistoricalAwards=false
• 关键词: NER; Biological; Nanomachines; Assembly; Function

Through evolution, biology has produced a remarkably diverse and efficient collection of proteins. These proteins assemble into a seemingly infinite variety of nanobiostructures, and they promote an extensive repertoire of chemical processes; hence, they comprise an interesting collection of biomolecular nanomachines. The proteins that are responsible for the maintenance and manipulation of DNA are one important subset of this collection. Their capacity to assemble with, and to alter the structure of, DNA is essential for all biological function. These proteins function in the packaging of DNA, the high-fidelity copying of genetic information, the reading of the genetic code and its conversion into RNA, the generation of genetic diversity, and the preservation of genetic and structural integrity of the genome. This project describes a new experimental approach to study the assembly and function of several of the nanomachines that function in these biological processes. The method involves the direct visualization by fluorescence microscopy, in real-time, of the assembly, disassembly, and movement of these nanobiostructures on single, optically-trapped DNA molecules using a novel, multi-port, laminar-flow, micro flow cell. This instrument will allows one to readily introduce an individual, optically-trapped DNA molecule sequentially into a series of reaction conditions, and to visualize the changes in structure/assembly of the molecules in real-time using multi-wavelength fluorescence microscopy. The successful development of this instrument will reveal important information about the structure and function of DNA-protein interactions that cannot be obtained using large ensembles of DNA molecules (where such information is often lost by averaging, or obscured by competing intermolecular interactions). This research will provide revolutionary new information about how proteins function to alter the structure of DNA. Furthermore, the experimental techniques can be applied to the study of many other nanoscale biostructures.

通过进化，生物学已经产生了非常多样化和有效的蛋白质集合。这些蛋白质组装成看似无限多样的纳米生物结构，它们促进了广泛的化学过程;因此，它们构成了生物分子纳米机器的有趣集合。负责维持和操纵DNA的蛋白质是这一集合的一个重要子集。它们与DNA组装并改变DNA结构的能力对于所有生物功能都是必不可少的。 这些蛋白质在DNA的包装、遗传信息的高保真复制、遗传密码的阅读及其转化为RNA、遗传多样性的产生以及基因组的遗传和结构完整性的保存中发挥作用。 该项目描述了一种新的实验方法来研究在这些生物过程中发挥作用的几种纳米机器的组装和功能。 该方法涉及通过荧光显微镜实时直接可视化这些纳米生物结构在单个光学捕获的DNA分子上的组装、拆卸和移动，使用新型的多端口层流微流动池。 该仪器将允许人们容易地将单个光学捕获的DNA分子顺序引入一系列反应条件中，并使用多波长荧光显微镜实时观察分子结构/组装的变化。该仪器的成功开发将揭示DNA-蛋白质相互作用的结构和功能的重要信息，这些信息不能使用大的DNA分子集合获得（这些信息通常因平均而丢失，或因竞争性分子间相互作用而模糊）。这项研究将为蛋白质如何改变DNA结构提供革命性的新信息。此外，实验技术可以应用于许多其他纳米生物结构的研究。